DESCRIPTION (applicant's description): We want to understand the molecular mechanisms responsible for motility and polarity in the budding yeast Saccharomyces cerevisiae and how these mechanisms interact with cell cycle control mechanisms. We are interested in how microtubules and their motors accomplish mitosis, in particular how the mitotic spindle is positioned based on interactions of cytoplasmic microtubules with the cell cortex. We studied how microtubules interact with the cortex during mitosis. We discovered evidence that microtubule sliding along the cortex is the mechanism by which dynein and dynactin move the spindle into the neck between mother and bud. We propose experiments to elucidate the molecular mechanisms that underlie microtubule sliding, especially how dynein, dynactin and microtubules interact with the cortex. We discovered evidence for a novel cell cycle checkpoint, in which cell division depends on proper positioning of the mitotic spindle. This checkpoint depends on the protein EBI. Our studies on how EB1 functions in cells suggest hypotheses for how the checkpoint is activated. We propose experiments to differentiate among these hypotheses. Also, we propose experiments aimed at understanding how cell division is delayed when the checkpoint is activated. We are interested in how the elements of the actin cytoskeleton function in polarization and transport. Cortical actin patches are polarized; we propose to track and quantitatively analyze the motion of patches to distinguish between models for patch polarization. We discovered that cytoplasmic actin cables are highly motile, by developing the ability to image actin cables in living cells. We will use this ability to investigate whether patch and cable movement depend on each other and if cables transport vesicles and RNA